Method and apparatus for generating a three-dimensional topographical image of a microscopic specimen

ABSTRACT

A method and apparatus for generating an image of a specimen comprises dividing the in-focus plane of an objective of an optical apparatus into a plurality of grid regions. Incident light is directed to the grid regions to illuminate surface portions of the specimen that are within the grid regions. Light reflected from the surface portions is sensed to determine approximate spatial slopes of each surface portion associated with each grid region. The spatial slopes are integrated to generate a topographical map representative of surface portions of the specimen that are within the depth of focus of the objective. Consecutive planes can be obtained to generate topographical maps thicker than the depth of focus of the objective.

BACKGROUND OF THE INVENTION

This invention relates to an apparatus and method of generating athree-dimensional topographic map (or image) of the surface of amicroscopic specimen, and for determining spatial slopes of surfaceportions of the specimen.

Several methods and devices have been employed and/or proposed fordetermining surface profiles of a microscopic specimen. One such methodemploys a reflected-light scanning profilometer that uses two pin-holeapertures and two light detectors, one behind each pin-hole aperture.The pin-hole apertures are located adjacent, but not exactly at,conjugate image planes. The apertures are slightly out of focus by thesame distance but in opposite directions. If the reflecting tile (i.e.portion of the specimen being observed) is in focus, the two detectorsrecord the same light intensity. If the specimen is moved away fromfocus, the intensity at one detector increases while the intensity atthe other decreases. The difference between the signals from the twodetectors is used in a feedback control loop that translates the stageuntil the reflecting tile is in focus. This method gives the location ofthe object tile in the z-direction, but fails to consider the effects ofthe tilt angle of the reflecting tile.

Another method proposes a confocal scanning profilometer having two slitapertures which are near conjugate image planes but slightly out offocus. This method recognizes that dual out-of-focus pin-hole aperturedesigns do not work when the reflecting tile has large tilt angles. Byusing slit apertures, this proposed method extends the range of tiltangles that may be collected. However, this method is also capable ofproviding only the location of the object tile in the z-direction; it isincapable of determining the spatial slope (i.e., the slope consideredwith relation to space or three-dimensional slope) of the reflectingtile.

Another type of device is an interference (heterodyne) profilometer.With this type of device, two mutually coherent laser beams of slightlydifferent frequency are combined or superimposed. The resulting beam hasa beat at a frequency that may be recorded by electronic instruments.The combined beam is split into a reference beam and a probing beam.Depth information is obtained from the phase difference between thereference beam and the probing beam. These instruments measure depth,not slope. Moreover, microphonics (i.e., acoustic noise) may alter theoptical path length of the two beams relative to one another therebyresulting in wrong measurements. Another potential difficulty is thermalexpansion, which may also alter the optical path lengths of the twobeams relative to one another.

To avoid or ameliorate measurement errors introduced by microphonics andthermal expansion, the probing beam and the reference beam may be madeto travel through a common path. This may be accomplished by splitting alaser beam into two components, one of which is frequency-shifted by anacoustic wave. After frequency-shifting, the two beams are superimposed.The resulting beam illuminates the back focal plane of an objective thathas a hole in the middle. The portion passing through this hole servesas a reference beam. The portion that passes through the glass in theobjective focuses into a diffraction-limited spot and serves as theprobing beam. Both portions are reflected and propagate back through acommon path up to a beam splitter before they are detected. On the lightreturn path, the two beams are concentric. The reference beam travels inthe center of the probing beam but without overlapping. Thus the twobeams share a common path and the instrument is less affected bymicrophonics and thermal expansion. However, this instrument isincapable of measuring the spatial slope of a reflecting tile.

Another method is based on projecting a sinusoidal grating into thespecimen. It records the reflected image for different phase angles ofthe projected grating. This results in a series of intensities for eachimage pixel. The phase of the discrete Fourier transform of the seriesof intensity is used to determine the phase of the grating reflected offthe specimen relative to that reflected off a reference planeperpendicular to the optical axis. The phase difference is then used todetermine height. This method does not provide angular information orotherwise determine the spatial slope of portions of the specimen.

Another method employs a confocal scanning optical profilometer. Thisdevice employs a dual-pin-hole aperture/dual detector arrangement thathas an aperture on a confocal back focal plane in front of the pin-holeaperture. This device is capable of detecting tilt angles in only oneorientation. Moreover, because this device uses two optical paths andtwo detectors, a tedious alignment and calibration procedure isnecessary because the two optical paths are not identical and thedetectors do not necessarily have identical sensitivity to light.

SUMMARY OF THE INVENTION

Among the several objects of the present invention is the provision ofan apparatus and method employing a confocal microscope for obtaininghigh resolution topographies of surfaces that reflect light; theprovision of an apparatus and method for determining spatial slopes ofsurface portions (e.g., reflecting tiles) of a microscopic specimen; theprovision of such an apparatus and method which is capable ofdetermining such spatial slopes even when such slopes are relativelylarge; the provision of such an apparatus and method which does notdepend on phase or coherent light; the provision of such an apparatusand method in which microphonics and thermal expansion have little or noeffect; the provision of such an apparatus and method which does notrequire tedious calibration and alignment procedures; the provision ofan apparatus and method for generating a map (or image) of the surfaceof the specimen; and the provision of such an apparatus and method inwhich an image may be generated relatively quickly and accurately.

In general, a method of the present invention for generating an image ofa specimen comprises dividing the in-focus plane of an objective of aconfocal microscope into a plurality of grid regions. Incident light isdirected to the grid regions to illuminate surface portions of thespecimen that are within the grid regions. Light reflected from thesurface portions is sensed to determine approximate (i.e.,representative) spatial slopes of each surface portion associated witheach grid region. The spatial slopes are integrated to generate atopographical map representative of surface portions of the specimenthat are within the depth of focus of the objective.

In another aspect of the present invention, a method of generating animage of a specimen comprises dividing the in-focus plane of anobjective of a confocal microscope into a plurality of grid regions. Thespecimen is positioned a distance from the objective of the opticalapparatus so that surface portions of the specimen are within the gridregions of the in-focus plane. Incident light is directed to the gridregions to illuminate the surface portions of the specimen that arewithin the grid regions. Light reflected from said surface portions issensed to determine approximate spatial slopes of each surface portionassociated with each grid region. The distance between the specimen andobjective is then altered so that other surface portions of thespecimen, which were previously outside the grid regions of the focalplane (e.g., outside the depth of field of the objective lens), arewithin previously unoccupied grid regions of the in-focus plane.Incident light is directed to the grid regions to illuminate the othersurface portions of the specimen that are within the previouslyunoccupied grid regions. Light reflected from the other surface portionsis sensed to determine approximate spatial slopes of each of the othersurface portions associated with each of the previously unoccupied gridregions. The determined spatial slopes are then integrated to generate atopographical map representative of surface portions of the specimen.

In another aspect of the present invention, a method of approximating aspatial slope (i.e., determining a representative spatial slope) of asurface portion of a specimen comprises directing incident light alongan incident light path of a confocal microscope to the surface portionof the specimen to reflect light from the surface portion. An intensityprofile associated with the reflected light is sensed.

In another aspect of the present invention, a confocal microscope isconfigured for approximating a spatial slope of a surface portion of aspecimen. The microscope has optical elements, an exit pupil, and asensor. The optical elements are configured for directing incident lightalong an incident light path from a light source to the surface portionof the specimen and for directing light reflected from the surfaceportion along a return light path. The exit pupil is in the return lightpath and is positioned and configured so that the exit pupil isilluminated by the reflected light in a manner (e.g. profile) dependentupon the spatial slope of the surface portion of the specimen. A sensoris in the return light path for generating at least one signalrepresentative of the profile of illumination of the exit pupil.

In yet another aspect of the present invention, a confocal microscope isconfigured for generating an image of a specimen. The microscope has anobjective, optical elements, a sensor, and a processor. The objective isconfigured for being spaced a distance from the specimen at which atleast part of the specimen is within the in-focus plane of theobjective. The optical elements are configured for directing incidentlight to discrete regions of the in-focus plane of the objective toilluminate surface portions of the specimen that are within of thediscrete regions. The sensor is configured for sensing light reflectedfrom the surface portions and for generating signals representative ofthe sensed light. The processor is configured for approximating spatialslopes of each surface portion of the specimen within the discreteregions of the in-focus plane.

Other objects and features will be in part apparent and in part pointedout hereinafter.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a schematic of a reflected light confocal scanning microscopeof the present invention showing incident light being focused on anobject tile of a specimen;

FIG. 2 is a schematic of the microscope of FIG. 1 showing a return pathof light reflected off the object tile;

FIG. 3 shows a projection of the reflected light beam at the back focalplane of the objective of the microscope of FIG. 1, the projectionrepresenting a portion of the objective's exit pupil illuminated by thereflected light beam;

FIG. 4 is a front view of a moving aperture (i.e., masking element) ofthe microscope of FIG. 1;

FIG. 5 represents the projection of the reflected light beam effectivelydivided into six area portions by rotating the masking element of FIG. 4to six discrete positions, each area portion corresponding to an area ofa conjugate back focal plane which is common to the aperture of themasking element for a given rotational position of the masking element;

FIG. 6 is an intensity versus position diagram which represents thelight intensity of the projected light beam common to each of the sixarea portions of FIG. 5. The intensity distribution is determined by thetilt, orientation, and numeric aperture of the objective;

FIG. 7 is a perspective view showing the geometrical relationshipbetween the incident and reflected light for the microscope of FIG. 1;and

FIG. 8 is a drawing of an image island showing the fifteen non-trivialclasses of pixels.

Corresponding reference characters indicate corresponding partsthroughout the several views of the drawings.

DESCRIPTION OF THE PREFERRED EMBODIMENTS

Referring now to the drawings, and first more particularly to FIG. 1, areflected light confocal scanning profilometer or microscope of thepresent invention is indicated in its entirety by the reference numeral20. The microscope is used for obtaining high resolution topographies oflight reflecting surfaces of a specimen (whether organic or inorganic).

Generally speaking, the microscope analyzes thousands of minusculeportions of the surface of a specimen. Because these surface portionsare so small relative to the size of the specimen, they are treated asthough they are flat. The apparatus (microscope and processor)determines the approximate (representative) spatial slopes of thesurface portions (or object tiles). A processor integrates thedetermined spatial slopes to generate a topographical map representativeof surface portions of the specimen. For simplicity, only one objecttile 22 is shown in FIGS. 1 and 2 and this tile appears to be largerelative to the components of the microscope 20. However, it is to beunderstood that the object tile 22 is only one of perhaps thousands ofobject tiles of the specimen, and each object tile is microscopic.

The microscope 20 is shown schematically in FIGS. 1 and 2. It comprisesa scanning/de-scanning mechanism 24, a beam splitter 26, an objective28, a moving aperture 30, a relay lens 32, a confocal pinhole aperture34, a light detector 36, and a processor 38. FIG. 1 illustrates anincident light path and FIG. 2 illustrates a return light path. Forsimplification in illustrating the return light path, thescanning/de-scanning mechanism 24 and beam splitter 26 have been omittedfrom FIG. 2. However, it is to be understood that thescanning/de-scanning mechanism 24 and beam splitter 26 are presentduring operation of the microscope 20.

Referring now to FIG. 1, incident light 40 is directed along theincident light path to the specimen via the beam splitter 26,scanning/de-scanning mechanism 24, beam splitter 26 and objective 28.Preferably, the scanning/de-scanning mechanism 24 comprises a rasterscan (not shown) and suitable lenses (not shown) for serially directinga plurality of collimated incident light beams off the beam splitter 26and through the objective 28 to serially illuminate different surfaceportions (object tiles) of the specimen. The objective 28 focuses theincident light beams onto the object tiles. The light beams (only one ofwhich is shown in FIG. 1) emitted from the scanning/de-scanningmechanism 24 are directed to the objective 28 at different angles sothat the beams are focused at different portions of the in-focus plane42 of the objective 28. In other words, the scanning/de-scanningmechanism 24 serially directs incident light to a plurality of discreteregions of the in-focus plane 42 of the objective 28. In essence, thescanning/de-scanning mechanism 24 divides the in-focus plane 42 of theobjective 28 into a plurality of grid regions 45 (e.g., 512×512 gridregions) and serially directs incident light to each grid region (note:grid regions and object tile are greatly enlarged in FIG. 1). If anobject tile 22 of the specimen is in a grid region of the in-focus plane42, then the incident light reflects off the object tile. Although thein-focus plane 42 is identified as a plane, it is to be understood thatit actually has a thickness equal to the depth of focus of the objective28. Likewise, each grid region has a thickness t (i.e., a distance fromleft to right as viewed in FIG. 1) preferably equal to the depth offocus of the objective 28.

The approximate spatial slope of an object tile 22 is determined byanalyzing the light beam reflected off such object tile. As shown inFIG. 2, the reflected light beam 44 is transmitted back through theobjective 28, through the beam splitter 26, through the relay lens 32,through the confocal pinhole aperture 34 and to the light detector 36.The incident light beam 40 (FIG. 1) focused onto each object tile 22 hasa particular shape. Preferably, the incident light beam is in the shapeof a right-circular cone having its base at the objective 28, its apexat the in-focus plane, and its axis generally aligned with the opticalaxis. Because of the laws of reflection, the reflected light beam 44will also be in the shape of a right-circular cone, but the position ofits axis will depend on the spatial slope of the object tile 22. Ingeneral, the axis of the reflected light beam 44 will be at an angleequal to twice the tilt angle of the object tile 22. The reflected lightwill be projected onto the back focal plane 46 of the objective 28(i.e., will illuminate a portion of the objective's exit pupil 48) onlyif the tilt angle is less than the NA (numerical aperture) angle of theobjective 28. If the tilt angle is greater than the NA angle, then noneof the reflected cone will return through the objective 28. Preferablythe objective 28 has a large NA angle (e.g., 50° or larger) to maximizethe range of slopes that can be detected by the microscope. FIG. 3 showsa projection 50 of the reflected light beam at the back focal plane 46of the objective 28. This projection 50 represents the portion of theexit pupil 48 illuminated by the reflected beam 44 of light. Since thereflected light beam 44 is cone-shaped, the projection 50 will be aconic section (i.e., circle, ellipse, parabola, or hyperbola). Theprecise shape and position (i.e., orientation) of the projection 50relative to the periphery of the exit pupil 48 is dependent upon thespatial slope of the object tile 22. As discussed in greater detailbelow, once the shape and position of the projection 50 is determined(or when characteristics of this shape and position are determined),then the spatial slope of the object tile 22 can be determined.

Preferably, characteristics of the shape and intensity of the light beamprojection 50 sufficient to determine the spatial slope of the objecttile are sensed by a single light detector, such as a single photodiode.A photodiode generally senses only the intensity of light striking thephotodiode and not the orientation of the light striking the photodiode.However, by placing a masking element in a plane 51 (FIG. 2) conjugateto the back focal plane 46, the light intensity of different portions ofthe back focal plane can be analyzed.

Referring now to FIGS. 2 and 4, the moving aperture 30 (i.e., maskingelement) is preferably positioned in a plane 51 conjugate to the backfocal plane 46 of the objective 28. Preferably, it includes adisc-shaped body 52 rotatable about the optical axis, and an offsetaperture 54 through the disc shaped body. The disc-shaped body 52 ispreferably opaque and the aperture 54 is preferably wedge-shaped. Thedisc-shaped body 52 masks (e.g., blocks) reflected light which wouldotherwise strike the light detector 36. Thus, only the portion of thereflected light beam 44 passing through the offset aperture 54 ispermitted to strike the detector 36. The masking element 30 is rotatedto move the aperture 54 to a plurality of positions (i.e., angularorientations) and the intensity of the portion of the reflected lightbeam passing through the aperture 54 is recorded at each of thesepositions. Preferably, the wedge angle of the offset aperture 54 is 60°and six different images are collected with the aperture atnon-overlapping rotational positions. FIG. 5 represents the image of theback focal plane divided by radial lines 56 into six area portions 58a,58b, 58c, 58d, 58e, 58f. Preferably, the diameter of the circle swept bythe aperture 54 is at least as large as the diameter of the image of theexit pupil at the conjugate back focal plane 51. Each area portioncorresponds to an area of the conjugate back focal plane 51 which iscommon to the aperture 54 of the masking element 30 for a given positionof the masking element. For each of the six positions of the maskingelement 30, the detector 36 detects the light intensity of a projectionportion 60a, 60b, 60c, 60d, 60e, or 60f (i.e., the portion of theprojection 50 which is common to the corresponding area portion) andgenerates a light intensity signal 62a, 62b, 62c, 62d, 62f, or 62grepresentative of the light intensity of the projection portion. The sixlight intensity signals 62a, 62b, 62c, 62d, 62f, 62g associated with thesix area portions 58a, 58b, 58c, 58d, 58e, 58f of FIG. 5 are depictedgraphically in FIG. 6. The combined light intensity signals arerepresentative of an intensity profile, e.g., the shape and position ofthe illuminated portion of the pupil of the objective.

Although the wedge angle of the pie-shaped aperture 54 is preferably60°, it is to be understood that the aperture may have some other wedgeangle, e.g. 180°, or some other shape without departing from the scopeof this invention. It is also to be understood that different numbers ofrotational positions of the masking element 30 could be employed withoutdeparting from the scope of this invention. Depending upon the shape andsize of the aperture 54 and the number of different rotationalpositions, the area portions may overlap and/or may not combine to equalthe area of the exit pupil. For example, if the aperture has a wedgeangle of 180°, and the masking element is rotated to six positionsspaced at 60° increments, then there will be significant overlap. Also,if the aperture is a pie-shaped wedge having a wedge angle of 30° andthe masking element is rotated to six positions spaced at 60°increments, then the combined data collected by the detector 36 willrepresent only a fraction of the light intensity of the projection 50.However, this data is still sufficient to determine the approximatespatial slope of the object tile 22.

Preferably, a light intensity signal is collected on each of the objecttiles before the aperture 54 is moved to another position. Thus, theaperture is required to be in each of its six positions only once toobtain the needed data on all of the object tiles in the grid regions.Preferably, the aperture 54 is moved to each of its positions via asuitable stepper motor (not shown).

A CCD, multiple photomultiplier tubes (PMT's), an image tube, or anintensifier may alternatively be used to obtain information about thestructure of the exit pupil or of the conjugate back focal plane. Theinformation obtained is essentially the same as when moving the detectorwith the advantage that no moving parts are necessary.

The light coming through the pinhole aperture that reaches the PMT haswithin it the spatial information to create a topographic map of theoptical section being imaged. However, a single-PMT detector integratesall the incoming light. Adding a masking element in front of thedetector that blocks all the light except for that reaching the detectorfrom one direction (e.g. a pie-shaped wedge aperture) allows thedetector to sample only the light from one orientation. By rotating themasking element as the scan device remains illuminating one specimentile, the reflected light returning at a plurality of orientations maybe sampled. As the rotation takes place the light intensity will vary inseveral ways. In the manner described above, these profiles (a stream ofintensities as the aperture rotates) can be easily used to assess theorientation of the tile being illuminated. Thus, a rapid rotation of apie-shaped aperture will give a data stream that can be analyzed todetermine orientation and tilt angle. Other rotating aperture geometrieswith subtle advantages and disadvantages are also possible (e.g.parallel slit, half circle).

The mask may be placed in front of or behind the confocal aperture, butbecause diffraction at the confocal pinhole could potentially degradethe structure of the exit pupil, the preferred implementation samplesthe structure of the exit pupil at a plane in front of the confocalpinhole.

In either approach the number of detectors in the array does notdetermine the resolution of the resultant data. It is possible to obtaininformation about the tile tilt and orientation from only a fewdetectors or detector positions using computational methods.

Multiple confocal aperture scanning, such as the rotating disk scanningmicroscopes, may also be used for profilometry by placing the additionalmasking element in a plane conjugate to the back focal plane that isonly on the return-light path behind the confocal apertures. Because thelight from several points in the specimen is collected at the same time,this approach requires additional optics to form an image of thescanning disk into a spatially resolved detector (CCD camera, videocamera, etc.). In this way the contributions from the different scanningspots are detected separately. This implementation has the advantage ofpermitting the use of a conventional mercury arc lamp for illuminationinstead of a laser. Also, diffraction at the pinholes in the lightreturn path may be negligible.

It is to be understood that the above procedure for collecting theangular information may also be used with a multiple-aperture tandemscanning microscope (TSM) (not shown), in which the illumination anddetection apertures are physically different. The confocal exit pupilmask is placed at a conjugate back focal plane behind the confocalapertures in the detection path. As with the tandem scanning microscopewith the conjugate back focal plane-mask in front of the confocalapertures, a spatially resolved detector is placed either right behindthe detection confocal apertures or at a plane conjugate to the plane ofthese apertures.

After the light intensity signals are collected for the object tiles ofthe specimen in the grid regions, the distance between the specimen andobjective 28 are altered so that other surface portions of the specimen,which were previously outside the grid regions of the focal plane (i.e.,previously outside the depth of focus of the objective) are broughtwithin previously unoccupied grid regions of the in-focus plane. Thelight intensity signals are then again collected for the object tiles inthe grid regions to obtain the necessary information on these newlyoccupied grid regions. The distance between the specimen and objective28 is preferably altered to a sufficient degree and to a sufficientnumber of positions to obtain the necessary information for all surfaceportions of the specimen.

After the data are collected (i.e., after the light intensity signalsare collected), the relationship between the spatial slope of thesurface at the scanning spot and the measured data may be determinedbased either on optical principles or on measurements taken from acalibrated sample. This relationship may have the form of a mathematicalexpression or of a table of angles (or slopes) versus measurements. Ifthe relationship is a mathematical expression, its analytic or numericalsolution may be attempted. If the relationship is in the form of atable, its inversion may be accomplished by scanning the table to findthe entry that best matches the measured quantities. It may also beinverted by using the measured quantities, properly scaled, as indicesto a table. Another possible approach to invert the information from atable is to fit mathematical expressions for tilt and orientation asfunctions of the quantities to measure, then apply this expression tothe recorded or measured quantities.

For simplicity, only the method of forming a table based on opticalprinciples is discussed in detail below. However, it is to be understoodthat the other methods for determining the slope of the object tile maybe employed without departing from the scope of this invention.

Referring now to FIG. 7, the relation between the incident and reflectedrays is determined in a straightforward way using the laws of reflectionand refraction. For simplicity, most of the components of the microscope20 have been excluded from FIG. 7. When a scanning beam (i.e., incidentlight beam 40) illuminates the exit pupil ("ExP") 48 of the objective28, all of the illumination rays that pass through the exit pupil focusinto a spot in the reflecting specimen or object tile 22. In objectspace (i.e., on the specimen side of the objective) each ray has a tiltangle (γ_(i)) and an orientation angle (φ_(i)) that depend on theportion of the exit pupil through which the ray passes. It is convenientto specify this location by its polar coordinates (ρ_(i), φ_(i)), whereρ₁ is the distance of the ray to the optical axis (or center of the exitpupil) normalized such that ρ=1 represents the rim of the exit pupil,and φ_(i) is the angular coordinate with respect to an arbitraryreference axis (e.g., the ξ axis shown in FIG. 7) in the back focalplane. In object space, the orientation angle of the ray is directlyrelated to this angular coordinate φ_(i). The tilt angle γ_(i) isrelated to the radial coordinate and is given by:

    γ.sub.1 =tan .sup.-1  ρ.sub.i tan (γ.sub.NA)!(1)

whereγ_(NA) =sin ¹ (NA/n), NA is the objective's numerical aperture, andn is the refractive index of the immersion medium (i.e., the mediumbetween the objective and the specimen). The tilt angle γ_(r) and theorientation angle φ_(r) of the reflected ray are dependent upon the tiltand orientation angles of the incident ray and of a normal n_(s) to thereflecting tile, as predicted by the law of reflection (i.e. the angleof incidence is equal to the angle of reflection). A unit vector k_(i)parallel to the illuminating ray is represented by:

k_(i) =(cos α_(i), cos β_(i), cos γ_(i))=-(cos φ_(i) sin γ_(i), sinφ_(i) sin γ_(i), cos γ_(i)). A unit vector n parallel to the normal ofthe reflecting surface is represented by:

n=(cos α_(s), cos β_(s), cos γ_(s))=(cos φ_(s) sin γ_(s), sin φ_(s) sinγ_(s), cos γ_(r)) A unit vector parallel to the reflected ray isrepresented by:

k_(r) =(cos α_(r), cos β_(r), cos γ_(r))=(cos φ_(r) sin γ_(r), sin φ_(r)sin γ_(r), cos γ_(r)). The negative sign for the incident indicates thatthe ray propagates in the opposite direction from the reflected ray.Using the law of reflection, the unit vector parallel to the reflectedray is:

    k.sub.r =k.sub.i -2(k.sub.i ·n)n,

where (k·n) is the dot product of k and n. Once k_(r) is known, the tiltangle γ_(r) can be obtained from the third component of this vector (cosγ_(r)) and the orientation φ_(r) angle from the expression: ##EQU1##Also, the radial coordinate of the point where the reflect ray crossesthe exit pupil is given by the expression: ##EQU2## The reflected raythus found propagates through the optical system to the conjugate backfocal plane aperture (i.e., the aperture 54 of the masking element 30),or equivalently, to a light detector that is physically or opticallytranslated at a conjugate back focal plane (CBFP). The conjugate backfocal plane aperture 54 may be located either in front or behind theconfocal pinhole aperture. It is clear from FIGS. 5 and 7 that for eachposition of the detector or the conjugate back focal plane aperture 54,only a subset of the reflected rays are collected. The subset dependsnot only on the position of the detector or of the aperture in theconjugate back focal plane, but also on the tilt and orientation anglesof the reflecting tile.

Diffraction may have a significant effect when the light propagatesthrough distances that are long compared to the size of the exit pupilof the objective or to the conjugate back focal plane aperture 54. Inthese cases the Fresnel diffraction approximations usually hold. Tocalculate the light distribution in the diffracted field it is necessaryto calculate the two-dimensional distribution of light intensity at theback focal plane or conjugate back focal plane of the objective. Thiscalculation has two parts. First, the shape of the illuminated portionof the back focal plane (i.e. the reflected image of the exit pupil)must be found. Second, the light intensity I_(r) (ξ_(r) η_(r)) at eachpoint (ξ_(r) η_(r)) in this illuminated portion must be determined.

As discussed above, the shape of the illuminated portion of the backfocal plane or conjugate back focal plane must be a conic sectionbecause the reflected beam is a cone with its axis not necessarilyparallel to the optical axis. The equation of this conic section can befound by first finding an equation that describes the reflected cone oflight, for example in the form ƒ(ξ, η, z)=0, and evaluating at the valueof z corresponding to the back focal plane. There are many ways toobtain the equation of the reflected cone of light. However, the linearalgebra approach is preferred because of the shorthand notation that itaffords.

The equation of a circular cone with its axis parallel to the z-axis(i.e., optical axis) and angle γ_(NA) is

    ξ.sup.2 +η.sup.2 -Z.sup.2 tan .sup.2 γ.sub.NA =0

which may be written in matrix notation as

    x.sup.T Qx=0

where x= ξ, η, z!^(T) is a point in three dimensional space, !^(T)denotes matrix transposition, and ##EQU3## The axis of symmetry of thereflected cone is the reflection of the chief ray (i.e., theillumination ray through the center of the exit pupil). From the law ofreflection this ray has director cosines (sin 2γ_(s) cos φ_(s), sin2γ_(s) sin φ_(s), cos 2γ_(s)) where (γ_(s), φ_(s)) are the tilt andorientation angles of the reflecting tile. The reflected cone may betreated as having its axis of symmetry along the axis of a rotatedcoordinate system: x= ξ γ z!^(T). The equation of the reflected cone inthese rotated coordinates is simply

    x.sup.T Qx=0                                               (3)

with Q given by Equation (2). The relation between the x and x vectorsis given by

    x=Rx                                                       (4)

where R is the coordinate rotation matrix given by ##EQU4## The equationof the reflected cone in the original coordinates obtained fromEquations (3) and (4) is:

    x.sup.T R.sup.T QRx=0.                                     (6)

To obtain the equation of the conic section formed by the intersectionof this cone and the back focal plane (i.e., the projection 50 of thecone onto the back focal plane), Equation (6) is evaluated at the valueof z corresponding to the back focal plane and then the resultingquadratic form is reduced to a two-dimensional quadratic in r=(ξ, η).Because the coordinates are normalized such that ρ=1 at the rim of theexit pupil 48, the back focal plane is at z=cot γ_(NA). Thissubstitution and subsequent reduction of order gives:

     r+Q.sub.11.sup.-1 Q.sub.12 Cot γ.sub.NA !.sup.T Q.sub.11  r+Q.sub.11.sup.-1 Q.sub.12 cot γ.sub.NA!=cot .sup.2 γ.sub.NA (Q.sub.12.sup.T Q.sub.11.sup.-1 Q.sub.12 -q.sub.33)       (7)

where Q₁₁, Q₁₂, and q₃₃ are a 2×2 symmetric matrix, a 2×1 matrix, orcolumn vector, and a scalar, respectively, that result from partitioningR^(T) QR as: ##EQU5##

The intensity of the illumination light I_(i) (ξ_(i), η_(i)) at a point(ξ_(i), η_(i))in the exit pupil and the intensity at the reflection(ξ_(r) η_(r)) of this point are not equal. The reason is that an areaelement dA_(i) =dA(ξ_(i), η_(i)) and its reflection dA_(r) =dA(ξ_(r),η_(r)) are not of the same size. Thus, the light that covers dA_(i) ofthe illumination, covers a different area dA_(r) of the reflected lightpath. The ratio of the reflected intensity to the illumination intensityis therefore the ratio dA_(i) /dA_(r). The relation between thelocations of the incident and reflected rays in the back focal plane aredenoted by ξ_(i) =ξ_(i) (ξ_(r), η_(r)) and η_(i) =η_(i) (ξ_(r), η_(r)).The ratio of areas is: ##EQU6## is the Jacobian of the relation betweenthe incident and reflected ray locations. The relations ξ_(i) =ξ_(i)(ξ_(r), η_(r)) and η_(i) =η_(i) (ξ_(r), η_(r)) may be found as describedabove. Thus the intensity of the reflected light in the back focal planeor conjugate back focal plane is: ##EQU7## To calculate the tilt andorientation angles (γ_(s), φ_(s)) of each object tile or pixel (x_(s),y_(s)) in the specimen, a series of measurements {I_(j) (x_(s), y_(s));j=0, 1, . . . , N_(d) -1} is collected by moving the detector 36 or theconjugate back focal plane aperture 54 to N_(d) different positions.Once the measurements have been collected, the theory developed abovemay be used to obtain the tilt and orientation angles (γ_(s), φ_(s)) foreach pixel (x_(s), y_(s)) in the specimen. This inverse problem may beaddressed in any one of several possible ways. Some of the methodsare: 1) finding two analytic expressions or functions φ_(r) =φ(I) andγ_(r) =Γ(I) for I={I₀, I₁, . . . , I_(Nd-1) } and simply substitutingthe recorded intensities; 2) Finding N_(d) analytic expressions orfunctions I_(j) =I_(j) (γ_(s), φ_(s)) that relate the N_(d) intensitiesto the recorded angles and solving them numerically for (γ_(s),φ_(s));3) finding calibration curves or look up tables (LUT), e.g., atable of the light intensity collected at each conjugate back focalplane aperture position or detector position for each combination oftilt and orientation angle of the reflecting tile; 4) findingapproximate expressions or functions φ_(r) =φ(I) and γ_(r) =Γ(I) forI={I₀, I₁, . . . , I_(Nd-1) } and substituting the recorded intensities.The look up table may be an auxiliary step in finding these twofunctions.

The look up table may be obtained from theoretical computation or fromexperimental measurement from a calibrated target.

In the experimental method, a reflecting surface whose tilt andorientation angles can be precisely controlled is set under themicroscope and then the intensity for each position of the detector orconjugate back focal plane aperture is recorded for a number ofcombinations of tilt and orientation angles.

In the theoretical approach, the laws of reflection and refraction anddiffraction theory are used to predict the intensities collected at eachposition of the detector or the conjugate back focal plane aperture.This calculation is done for a number of combinations of tilt andorientation angles of the reflecting tile. The look up table may becalculated from closed form expressions that relate the intensities ateach position of the detector or conjugate back focal plane aperture tothe tilt and orientation angles. Alternatively, it may be calculatednumerically in the following way:

i) partition the exit pupil into small area elements {Δa_(i) ; i=1, 2, .. . , N_(a) };

ii) assign a tilt angle γ_(s) and orientation angle φ_(s) to thereflecting tile;

iii) clear one accumulator {I_(j) (γ_(s),φ_(s));j=0, 1, . . . , N_(d)-1} for each one of the N_(d) positions of the detector or conjugateback focal plane aperture;

iv) for an area element Δa_(i), calculate the intensity I(ξ_(i), η_(i))of the illumination at a point (ξ_(i), η_(i)) within the area element;

v) calculate the tilt and orientation angles γ_(i), φ_(i) in objectspace for an illumination ray passing through (ξ_(i), η_(i)) in the exitpupil;

vi) with (γ_(i), φ_(i)) and (γ_(s), φ_(s)), using the law of reflectioncalculate the tilt and orientation angles γ_(r), Φ_(r) of the reflectedsurface;

vii) with (γ_(r), φ_(r)), find the position(s) of the detector orconjugate back focal plane aperture where the reflected rays aredetected;

viii) Add Δa_(i) I(ξ_(i), η_(i)) to the intensity detected at theposition(s) found in Step vii;

ix) repeat steps iv to viii for all area elements {Δa_(i) ; i=1, 2, . .. , N_(a) } in the exit pupil;

x) if the intensities {I_(j) (γ_(s), φ_(s)); j=0, 1, . . . , N_(d) -1}computed in steps iv to ix are not all equal to zero, normalize the setof intensities to add to 1.0 (if all N_(d) intensities are zero, thetilt angle γ_(s) of the reflecting surface is larger than or equal tothe maximum angle collected by the objective, γ_(NA) and none of thereflected rays pass through the exit pupil); and

xi) repeat Steps ii to x for all angles 0≦φ_(s), <2π and0≦γ_(s) <γ_(NA).

The result of this process is a three-dimensional array of intensities:

    {I.sub.j (γ.sub.s,φ.sub.s);

j=0, 1, . . . , N_(d) -1, 0≦φ_(s),<2π, 0≦γ_(s) <γ_(NA) }

normalized such that ##EQU8##

Some geometries for the conjugate back focal plane aperture (or detectoraperture) allow for simplifications in the calculation of the look uptable, in particular when the illumination intensity is circularlysymmetric, i.e. I(ξ, η)=I(ρ), with ρ² =ξ² +η². If illumination iscircularly symmetric and the conjugate back focal plane aperture iswedge-shaped with angle θ_(w), i.e. for φ₀ ≦φ_(s) <φ₀ +θ_(w) for somearbitrary value of φ₀, then the intensities I_(j) (ξ_(s), η_(s)) for therest of the 0, 2, π) interval are rotations of the intensities in theφ₀, φ₀ +θ_(w)) interval. In other words, the N_(d) intensities {I_(j)(φ_(s), γ_(s)); j=0, 1, . . . , N_(d) -1}, for an orientation angleφ_(s) .epsilon slash. φ₀, φ₀ +θ_(w)) can be found from the intensitiesin the φ_(o), φ₀ +θ_(w) ! interval by some rotation of the apertureposition index j and orientation angle as:

    I.sub.j (φ.sub.s, γ.sub.s)=I.sub.(j+k)mod N.sbsb.d (φ.sub.s +α, γ.sub.s)                                  (9)

for some k and α that depend on θ_(w), φ₀, and possibly on N_(d), butnot on φ_(s) or γ_(s). In Equation (9) the angle φ_(s) +α is in theinterval φ₀, φ₀ +θ_(w)).

In practice, the optical path lengths between the different componentsin the optical system may be long compared to the size of the aperturesinvolved. For example, the distance from the objective 28 to the pinholeaperture 34 could be at least an order of magnitude larger than thediameter of the exit pupil of the objective. In these cases diffractionhas a significant effect on the distribution of light at differentplanes of interest. In the embodiment of FIGS. 1 and 2, the maskingelement (i.e., moving aperture 30) is at a conjugate back focal plane infront of the confocal pinhole aperture 34. In this embodiment thedistance from the objective 28 to the moving aperture 30 is severaltimes larger than the diameter of the objective's exit pupil 48 to allowenough room for the scanning/de-scanning mechanism 24 and the beamsplitter 26 that bring the illumination into the objective's back focalplane 46. Also, the distance from the relay lens 32 to the pinholeaperture 34 is large compared to the diameter of the exit pupil of theobjective 28 and relay lens 32. In such cases diffraction effects haveto be taken into account to calculate the look up table. In thecalculation proposed here, it is assumed that the tilt and orientationangles of the reflecting specimen do not change significantly within thesize of the diffraction limited illumination spot. The look up table iscalculated as follows:

i) assign a tilt γ_(s) and orientation φ_(s) to the reflecting tile;

ii) using Equation (8), calculate the reflected light intensity I_(r)(ξ_(r), η_(r)) in the ExP;

iii) use diffraction theory to calculate the optical disturbance U_(c-)(ξ, η) in front of the conjugate back focal plane aperture;

iv) for a given position j of the conjugate back focal plane aperture,calculate the optical disturbance U_(c+) (ξ, η) behind the conjugateback focal plane aperture: U_(c+) (ξ, η)=U_(c-) (ξ, η) for (ξ, η) withinthe transparent part of the aperture, and U_(c+) (ξ, η)=0 for (ξ, η)within the opaque part of the aperture;

v) use diffraction theory to calculate the optical disturbance U_(L-)(ξ, η) in front of the relay lens 32;

vi) multiply U_(L-) (ξ, η) by the phase factor introduced by the lens toobtain the optical disturbance U_(L-) (ξ, η) behind the lens;

vii) use diffraction theory to calculate the optical disturbance U_(p-)(ξ, η) in front of the confocal pinhole aperture;

viii) multiply U_(p-) (ξ, η) by the confocal aperture pupil functionU_(c) (ξ, η) to obtain the optical disturbance U_(p+) (ξ, η) behind thepinhole aperture;

ix) integrate the intensity of the optical disturbance U_(p+) (ξ, η)behind the pinhole aperture and assign the intensity to I_(j)(φ_(s),γ_(s));

x) repeat Steps iv to ix for each position of the conjugate back focalplane aperture j=0, 1, . . . , N_(d) -1;

xi) if the intensities {I_(j) (γ_(s), Φ_(s)); j=0, 1, . . . N_(d) -1}computed in steps iv to x are not all equal to zero, normalize the setof intensities to add to 1.0;

xii) repeat steps i to xi for all angles 0≦φ_(s) <2π and 0≦γ_(s)<γ_(NA).

As with the geometrical optics approach the result is athree-dimensional array of intensities:

    {I.sub.j (γ.sub.s, φ.sub.s);

j=0, 1, . . . , N_(d) -1, 0≦Φ_(s) <2π, 0≦γ_(s) <γ_(NA) }

normalized such that ##EQU9## The simplifications afforded by certainconjugate back focal plane-aperture geometries and illuminationsymmetries for the geometric optics approach can also be exploited whendiffraction effects are important.

In an alternative embodiment (not shown), the moving aperture is at aconjugate back focal plane behind the confocal pinhole aperture. Thecalculation of the look up table is very similar to the one describedfor the case of a moving aperture in front of the confocal pinhole. Thatis, the light distribution U_(d) (ξ, η) in front of the light detectorhas to be calculated from the light distribution I_(r) (ξ, η) in theback focal plane of the objective taking into account the diffractioneffects from one plane of interest to the next. In this setup the sizeof the confocal pinhole aperture is an important design parameter. Ifthe confocal aperture is too large, it will not prevent out-of-focuslight from reaching the detector. If, on the other hand, it is toosmall, diffraction effects will distort the structure of the conjugateback focal plane to the point of practically losing the informationabout the tilt and orientation angles of the reflecting specimen.

After the look up table (LUT) is developed, it can be used to obtain thespatial slope (i.e., tilt and orientation) for each measured image. Forexample, the look up table may be scanned to find the value(s) closestto the measured intensity associated with each object tile.

The look up table may also be used to derive approximate expressions fortilt and orientation in terms of the intensities, i.e., to approximatetwo functions φ_(r) =Φ(I) and γ_(r) =Γ(I) for I={I_(o), I₁, . . . ,I_(Nd-1) }. Possible approaches for obtaining these functions include:(1) least squares fit of arbitrary functions (using singular valuedecomposition or not); (2) polynomial interpolation (or interpolationusing an arbitrary set of functions); and (3) Minimax (Chevyshev)theory.

After being calculated, the spatial slopes of the object tiles areintegrated to form a topographical map of the reflecting surface. Theslopes have to be calculated in at least two directions, preferablymutually orthogonal (e.g., horizontal and vertical directions). If theslopes at column i, row j of the image are ##EQU10## then, to afirst-order approximation, the height z at pixel (i, j) can becalculated either as

    Z.sub.i, j ≅Z.sub.i-1, j +Δxg.sub.i-1, j   (10)

or as

    Z.sub.i, j ≅Z.sub.i, j-1 +Δyh.sub.i, j-1   (11)

where Δx and Δy are the pixel sizes in the horizontal and verticaldirections. Because of the integrability problem, Equations (10) and(11) will seldom give the same value for z_(i), j. One cause of theintegrability problem is that Equations (10) and (11) are first orderapproximations that are exact only if the reflecting surface is a planeor if Δx and Δy approach zero. Another cause is that, even whenEquations (10) and (11) are exact, there is measurement noise and thusthe slopes g_(i), j and h_(i), j are never exact. Using only eitherEquation (10) or Equation (11), does not work around the integrabilityproblem. For example, suppose that Equation (10) is used to calculatethe surface height along two adjacent rows, j and j+1 each with N_(x)pixels. Each pixel height z_(i+1), j is calculated from its immediatelypreceding neighbor z_(i), j. If z_(i), j has some error, z_(i+1), j willhave the cumulative effects of this error and any error resulting fromEquation (10) (either from measurement error or from using a first orderapproximation). Likewise z_(i+2), j will have the cumulative effects oferrors in z_(i+1), j, z_(i), j, g_(i+1) j, and g_(i), j. This erroraccumulation propagates all across row j, and thus Z_(Nx), j will havethe accumulated error of all other pixels in the row. Likewise, Z_(Nx),j+1 will have the accumulated error of all other pixels in row j+1.Because the errors at each row are likely to be different and because ofthe cumulative effect of the errors, the final pixel heights in rows jand j+1, z_(Nx), j and z_(Nx), j+1, are likely to be significantlydifferent.

Another drawback of line-by-line integration of the slopes is that in atypical image there are a large number of dark pixels. These pixelscorrespond to portions of the specimen that either are out of focus orhave tilt angles larger than γ_(NA). Out-of-focus pixels are darkbecause most of the light reflected off these pixels is masked by theconfocal aperture. Pixels with tilt angles larger than γ_(NA) reflectall the light at angles larger than γ_(NA), and thus all the lightreflected off these pixels misses the objective's exit pupil. Theout-of-focus pixels may not be a problem if enough optical slices arecollected to include the whole depth of the specimen. Unfortunately,pixels with large tilt angles are out of the range of measurable slopesand they appear dark in all optical sections. At the dark pixels, thetilt and orientation information is missing and thus the assumption thateach pixel is connected to its next neighbor is violated. Line-by-lineintegration can be applied only if the image region is simply-connectedand "convex", i.e., if the image region does not have holes or gulfs.Even in those cases, adjacent lines can have similar topography only ifthe region is a two-dimensional interval, i.e. a rectangular region ofthe collected image. For an arbitrary simply-connected convex region,however, the height along one line is very likely to be significantlyoffset relative to the adjacent lines because they have differentstarting points.

To avoid the integrability problem and to be able to obtaintopographical maps of non-convex regions, the surface profile iscalculated using a least squares approach. For multiple-connectedregions, the topographical map of each one of the simply-connected imageregions or islands is calculated. For each island I_(n) the sum ofsquared errors is defined as ##EQU11## and a set of surface heights(z_(i), j) is found that makes E a minimum. To obtain this minimum wefollow the standard procedure of setting ##EQU12## and solving forz_(i), j. Equation (13) leads to a linear system of equations of theform

    Az=h                                                       (14)

where A is an N×N matrix of constant coefficients, z is a vectorcontaining the N surface heights z_(i), j in lexicographical ordering, his an N-element vector whose elements depend on g_(i), j Δx and h_(i), jΔy, and N is the number of pixels in the image island. The slopesprovide information about the height of a pixel relative to itsneighbors, thus in Equation (14) there is one pixel height that can bearbitrarily chosen.

Equation (14) can be solved using any one of a number of numericalmethods. For any practical image size, however, the number of pixels, N,is very large and the size of matrix A is impractically large. Forexample, a small 128×128 image, (i.e. an image with 128 rows and 128columns) N may be up to 16,384 pixels, and a typical 512×512 image mayhave as many as 262,144 pixels. However, A is a scarce matrix with atmost 5 non-zero elements per row. This is because in Equation (12) eachpixel z_(ij) depends only on its immediate neighbors z_(i)±1, j±1, andeach pixel can have at most four immediate neighbors (boundary pixelshave less than four neighbors). Because A is a scarce matrix, iterativemethods to solve Equation (14) have small per-iteration computationalcomplexity and thus such methods are chosen to solve this equation. Inparticular, the Gauss-Seidel iteration method is employed to solveEquation (14) for z. For Equation (12), this simply means that z_(i),j.sup.(k+1) is the average of all possible ways to calculate the heightat pixel (i, j) using a first order approximation given the heights ofits immediate neighbors at iteration k or (k+1). For example, if pixel(i, j) is an interior pixel, it has four immediate neighbors that areimage pixels, and thus there are four possible ways to calculate theheight of pixel (i, j), namely from the left, from the right, fromabove, and from below. This gives ##EQU13## Similar equations are easilyderived for other classes of pixels. These equations may be cast in thegeneral form ##EQU14## Implementation of Equation (16) as written aboveis inefficient for computational purposes due to the large number oftrivial multiplications (by zero or by 1, 0). A more efficientcomputation results if, before solving Equation (14), the pixels in animage island are classified according to the possible ways to calculatez_(i), j, that is according to the number and location of their non-darkimmediate neighbors. Then a different equation is used for each class ofpixel. Because each pixel in an island may have up to four non-darkneighbors, there are sixteen possible classes of pixels. From these,there is a trivial class that has zero non-dark neighbors because theisland consists of a single pixel. In this case, the pixel height cannotbe computed relative to any other pixel. For an isolated pixel, thedenominator in Equation (16) is exactly zero and computation of thepixel height is therefore not possible. The remaining fifteennon-trivial classes of pixels are shown in FIG. 8. Once the pixels in anisland are classified, the first pixel of the island is selected suchthat in a raster scan of all the pixels in the island, this will be thefirst pixel scanned. This is either an upper-tip pixel (See FIG. 8) oran upper-right corner pixel. Then Equation (16) is applied in a rasterscan for all pixels in the island. One visit to all pixels completes aniteration. The iteration is repeated until the change in pixel heightfrom one iteration to the next is arbitrarily small.

Even with the more efficient implementation used for Equation (16), theGauss-Seidel method has a slow global convergence rate. To accelerateits convergence rate we use the standard method of successive overrelaxation (SOR). Equation 16 thus becomes: ##EQU15## where ω is theoverrelaxation factor. For stability 0<ω<2. The SOR method acceleratesthe global convergence by an order of magnitude, however, the localconvergence rate varies greatly across the pixels in an island. Pixelsthat are close to the first pixel reach their final value much fasterthan pixels that are farther from this first pixel. To accelerate thelocal convergence rate, the direction of the raster scan is alternatedbetween iterations, by the alternating-direction iterative (ADI) method.In each iteration of this method the direction of the raster scan ischanged. In the first iteration the raster scan is performed line byline from left to right and top to bottom, the second iteration is alsoline by line but from right to left and bottom to top, in the thirditeration the raster scan is done column by column from top to bottomand from left to right, in the fourth iteration the raster scan is donealso column by column but from bottom to top and from right to left.This pattern of four raster scan directions is then repeated for allsubsequent iterations. Simplified versions of this method are possiblewhich use only two (or three) directions for the raster scan, however,the local convergence rate is faster with this four-directional ADIimplementation.

In view of the above, it will be seen that the several objects of theinvention are achieved and other advantageous results attained.

As various changes could be made in the above constructions and methodswithout departing from the scope of the invention, it is intended thatall matter contained in the above description or shown in theaccompanying drawings shall be interpreted as illustrative and not in alimiting sense. The invention therefore shall be limited solely by thescope of the claims set forth below.

What is claimed is:
 1. A method of generating an image of a specimencomprising:dividing the in-focus plane of an objective of a confocalmicroscope into a plurality of discrete regions; directing incidentlight to the discrete regions to illuminate surface portions of thespecimen that are within the discrete regions; sensing light reflectedfrom said surface portions; determining an approximate spatial slope ofeach surface portion associated with each discrete region; andintegrating said determined spatial slopes to generate a topographicalmap representative of surface portions of the specimen that are withinthe depth of focus of the objective.
 2. A method as set forth in claim 1wherein the discrete regions of the in-focus plane of the objective areof a thickness substantially equal to the depth of focus of theobjective.
 3. A method as set forth in claim 1 wherein the step ofdirecting incident light to the discrete regions comprises directingseparate beams of incident light to the regions so that said reflectedlight comprises separate beams, the beams of reflected lightcorresponding to the surface portions of the specimen.
 4. A method asset forth in claim 1 wherein the step of directing light to the discreteregions comprises serially directing light to each discrete region toseparately illuminate each surface portion within a corresponding one ofthe discrete regions.
 5. A method as set forth in claim 1 wherein theapproximate spatial slope of each surface portion is determinedby:generating at least one signal representative of an intensity profileassociated with light reflected off said each surface portion; andapplying known values to said at least one generated signal.
 6. A methodas set forth in claim 5 wherein applying known values to said at leastone generated signal comprises comparing said at least one generatedsignal to one of a plurality of known values, each known value beingassociated with light reflected from a surface of known spatial slope.7. A method as set forth in claim 1 wherein the step of sensingreflected light comprises sensing an intensity profile associated withthe shape and position of a projection of at least a portion of thereflected light onto a particular area of a given plane.
 8. A method asset forth in claim 7 wherein said particular area of said given planecomprises a plurality of area portions, and wherein sensing an intensityprofile associated with the shape and position of said projectioncomprises generating a plurality of signals, each of which isrepresentative of an area common to said projection and to one of saidplurality of area portions.
 9. A method as set forth in claim 8 whereinsaid plurality of signals are generated by a single light sensor.
 10. Amethod as set forth in claim 7 wherein sensing an intensity profileassociated with the shape and position of said projectioncomprises:positioning a masking element relative to said given-plane sothat a portion of said particular area of said given plane is masked andanother portion of said particular area is unmasked, said maskingelement being configured such that movement of the masking elementvaries the portion of said particular area which is unmasked; moving themasking element to a plurality of discrete positions relative to saidgiven plane, a plurality of unmasked area portions of said particulararea of said given plane corresponding to the plurality of discretepositions of the masking element; and generating a plurality of signals,each signal being representative of an area which is common to saidprojection and to the unmasked area portion associated with each of saidpositions of the masking element.
 11. A method of generating an image ofa specimen comprising:dividing the in-focus plane of an objective of aconfocal microscope into a plurality of grid regions; positioning thespecimen a distance from the objective of the optical apparatus so thatsurface portions of the specimen are within the grid regions of thein-focus plane; directing incident light to the grid regions toilluminate the surface portions of the specimen that are within the gridregions; sensing light reflected from said surface portions to determineapproximate spatial slopes of each surface portion associated with eachgrid region; altering the distance between the specimen and objective sothat other surface portions of the specimen, which were previouslyoutside the grid regions of the focal plane, are within previouslyunoccupied grid regions of the in-focus plane; directing incident lightto the grid regions to illuminate said other surface portions of thespecimen that are within said previously unoccupied grid regions;sensing light reflected from said other surface portions to determineapproximate spatial slopes of each of said other surface portionsassociated with each of said previously unoccupied grid regions; andintegrating said determined spatial slopes to generate a topographicalmap representative of surface portions of the specimen.
 12. A method asset forth in claim 11 wherein the approximate spatial slope of eachsurface portion is determined by:generating at least one signalrepresentative of a characteristic associated with light reflected offsaid each surface portion; and applying known values to said at leastone generated signal.
 13. A method as set forth in claim 12 whereinapplying known values to said at least one generated signal comprisescomparing said at least one generated signal to one of a plurality ofknown values, each known value being associated with light reflectedfrom a surface of known spatial slope.
 14. A method of approximating aspatial slope of a surface portion of a specimen comprising:directingincident light along an incident light path of a confocal microscope tosaid surface portion of the specimen to reflect light from said surfaceportion; sensing a characteristic associated with said reflected light;and applying the sensed characteristic to values relating to spatialslope.
 15. A method as set forth in claim 14 wherein sensing acharacteristic associated with said reflected light comprises sensing anintensity profile associated with said reflected light.
 16. A method asset forth in claim 15 wherein said values comprise a plurality of knowncharacteristics, each of said known characteristics being associatedwith reflection of light from a surface of known spatial slope, andwherein the step of applying the sensed characteristic comprisesmatching the sensed intensity profile to one of said knowncharacteristics.
 17. A method as set forth in claim 15 wherein the stepof directing light comprises directing incident light of a particularshape and direction to said surface portion so that said reflected lightis of a particular shape.
 18. A method as set forth in claim 17 whereinthe step of sensing an intensity profile associated with said reflectedlight comprises sensing an intensity profile associated with the shapeand position of a projection of at least a portion of the reflectedlight onto a particular area of a given plane.
 19. A method as set forthin claim 18 wherein said particular area of said given plane comprises aplurality of area portions, and wherein sensing an intensity profileassociated with the shape and position of said projection comprisesgenerating a plurality of signals, each of which is representative of anarea common to said projection and to one of said plurality of areaportions.
 20. A method as set forth in claim 19 wherein said pluralityof signals are generated by a single sensor.
 21. A method as set forthin claim 18 wherein sensing an intensity profile associated with theshape and position of said projection comprises:positioning a maskingelement relative to said given plane so that a portion of saidparticular area of said given plane is masked and another portion ofsaid particular area is unmasked, said masking element being configuredsuch that movement of the masking element varies the portion of saidparticular area which is unmasked; moving the masking element to aplurality of discrete positions relative to said given plane, aplurality of unmasked area portions of said particular area of saidgiven plane corresponding to the plurality of discrete positions of themasking element; and generating a plurality of signals, each signalbeing representative of an area which is common to said projection andto the unmasked area portion associated with each of said positions ofthe masking element.
 22. A method as set forth in claim 20 wherein thestep of applying the sensed characteristic to values relating to spatialslope comprises applying the generated signals to values relating tospatial slope.
 23. A method as set forth in claim 22 wherein said valuescomprise a plurality of known values, each known value being associatedwith reflection of light from a surface of known spatial slope, andwherein the step of applying the generated signals comprises matchingthe generated signals to one of said known values.
 24. A method ofgenerating a topographical image of a specimen, said method comprisingthe steps of:directing incident light through an objective of a confocalmicroscope onto said specimen to thereby illuminate a plurality ofsurface portions thereof, sensing light reflected from only thosesurface portions thereof that are within said objective's depth of fieldand numerical aperture, determining from said sensed reflected light aspatial slope for said surface portions, and integrating said determinedspatial slopes to thereby generate a topographical map representative ofsaid light reflecting surface portions, said topographical map therebycomprising said topographical image of said specimen.
 25. A method asset forth in claim 24 further comprising the step of re-positioning saidspecimen with respect to said objective, as required, to bring more ofthe surface portions of said specimen within the objective's depth offield.
 26. The method as set forth in claim 25 wherein the step ofdirecting incident light includes the step of scanning said specimenwith the incident light to thereby sequentially illuminate separatesurface portions of the specimen.
 27. The method as set forth in claim26 wherein the step of sensing the reflected light includes the step ofsensing a profile of said reflected light.
 28. The method as set forthin claim 27 wherein the step of sensing a profile includes the step ofmarking said reflected light from each of said surface portions with anaperture, and illuminating again each of said surface portions as saidaperture is re-positioned.
 29. The method as set forth in claim 28wherein the integrating step includes the step of normalizing the valuesof height of adjacent determined spatial slopes.
 30. The method as setforth in claim 27 wherein the step of determining the spatial slopeincludes the step of calculating a spatial slope from the sensed profileof each surface portion.
 31. The method as set forth in claim 30 whereinthe step of calculating a spatial slope includes the step of referringto a look up table.
 32. A confocal microscope for generating atopographical image of a specimen, said device comprising:an incidentlight source for directing incident light through an objective onto saidspecimen to thereby illuminate a plurality of surface portions thereof,a detector in a reflected light path for detecting light reflected fromonly those surface portions that are within the objective's depth offield a numerical aperture, and an electronic processor configured fordetermining a spatial slope for said surface portions, and integratingsaid determined spatial slopes into a topographical map to therebygenerate a topographical image of the specimen.
 33. A confocalmicroscope for approximating a spatial slope of a surface portion of aspecimen, said microscope having:optical elements configured fordirecting incident light along an incident light path from a lightsource to the surface portion of the specimen and for directing lightreflected from the surface portion along a return light path, one of theoptical elements including an exit pupil in the return light path, theoptical elements being positioned and configured so that the exit pupilis illuminated by the reflected light in a manner dependent upon thespatial slope of the surface portion of the specimen; a sensor in thereturn light path for generating at least one signal representative ofthe manner of illumination of the exit pupil; and a processor formatching the generated signal to a signal associated with lightreflected from a surface of a known spatial slope.
 34. A confocalmicroscope for approximating a spatial slope of a surface portion of aspecimen, said microscope having:optical elements configured fordirecting incident light along an incident light path from a lightsource to the surface portion of the specimen and for directing lightreflected from the surface portion along a return light path, one of theoptical elements including an exit pupil in the return light path, oneof the optical elements including an objective having a numericalaperture angle of at least approximately 50°, the optical elements beingpositioned and configured so that the exit pupil is illuminated by thereflected light in a manner dependent upon the spatial slope of thesurface portion of the specimen; and a sensor in the return light pathfor generating at least one signal representative of the manner ofillumination of the exit pupil.
 35. A confocal microscope forapproximating a slope and orientation of a surface portion of aspecimen, said microscope having:optical elements configured fordirecting incident light along an incident light path from a lightsource to the surface portion of the specimen and for directing lightreflected from the surface portion along a return light path; and meansfor detecting characteristics of the reflected light sufficient toapproximate the spatial slope of the surface portion of the specimen.36. A microscope as set forth in claim 35 wherein said optical elementsare configured for directing incident light of a particular shape anddirection to said surface portion so that the reflected light is of aparticular shape.
 37. A microscope as set forth in claim 36 wherein saiddetecting means comprises means for sensing a characteristic associatedwith the shape and position of a projection of at least a portion of thereflected light onto a particular area of a given plane.
 38. Amicroscope as set forth in claim 37 wherein said sensing means includesa masking element positioned relative to said given plane and configuredso that a portion of said particular area is masked by the maskingelement and another portion of said particular area is unmasked by themasking element, the masking element being moveable relative to saidparticular area for varying the portion of said particular area which isunmasked.
 39. A microscope as set forth in claim 30 wherein saiddetecting means further comprises means for generating a plurality ofsignals, each signal being representative of an area which is common tosaid projection and to the unmasked area portion associated with each ofa plurality of discrete positions of the masking element relative tosaid particular area.
 40. A confocal microscope for generating an imageof a specimen, said microscope having:an objective configured for beingspaced a distance from the specimen at which at least part of thespecimen is within the in-focus plane of the objective; optical elementsconfigured for directing incident light to discrete regions of thein-focus plane of the objective to illuminate surface portions of thespecimen that are within the discrete regions; a sensor for sensinglight reflected from the surface portions and for generating signalsrepresentative of the sensed light; and a processor configured forapproximating, in response to the signals generated by the sensor,spatial slopes of each surface portion of the specimen within thediscrete regions of the in-focus plane.
 41. A microscope as set forth inclaim 40 wherein the processor is further configured for integrating thedetermined spatial slopes to generate a topographical map representativeof surface portions of the specimen.
 42. A microscope as set forth inclaim 40 wherein said optical elements are configured for directingincident light of a particular shape and direction to said discreteregions so that the reflected light is of a particular shape.
 43. Amicroscope as set forth in claim 42 wherein said optical elements areconfigured for projecting at least a portion of the reflected light ontoa particular area of a given plane.
 44. A microscope as set forth inclaim 43 further comprising a masking element positioned relative tosaid given plane and configured so that a portion of said particulararea is masked by the masking element and another portion of saidparticular area is unmasked by the masking element, the masking elementbeing moveable relative to said particular area for varying the portionof said particular area which is unmasked.
 45. A microscope as set forthin claim 44 wherein the sensor is configured for generating a pluralityof signals, each signal being representative of an area which is commonto the projection and to the unmasked area portion associated with eachof a plurality of discrete positions of the masking element relative tosaid particular area.
 46. A microscope as set forth in claim 40 whereinthe objective has a numerical aperture angle of at least approximately50°.
 47. A microscope as set forth in claim 40 wherein the objective hasa numerical aperture angle of at least approximately 60°.